Oestrus refers to the phase when the female is sexually receptive (“in heat”). Under regulation of gonadotropic hormones, ovarian follicles mature and estrogen secretions exerts its influence. The female then exhibits sexually receptive behaviour, a situation that may be signalled by visible physiologic changes. Oestrous synchronisation is the process of targeting female mammals to come to heat and ovulate within a short time frame (usually 36 to 96 hours) so they can be inseminated at approximately the same time to save time and costs and generally stream-line the process. Synchronization of the oestrous cycle is economically advantageous and most commonly used in agricultural animals for example to decrease the costs for artificial insemination and maximise the efficiency and profitability of milk production.
Induction and/or synchronization of oestrous cycle is routinely achieved in most theria mammalian species, by supplying different combinations of hormones; luteolytic (prostaglandin F2α, PGF2α and its analogues) and progestative (progesterone and its analogues) factors are commonly used. Secretion of PGF2α is the event that provokes the regression of the corpus luteum in mammals, giving way to a follicular phase which culminates in oestrus behaviour and therefore, ovulation. Oestrus can be said to be the precursor of ovulation. Administration of exogenous PGF2α, or its analogues to livestock animals such as pigs, cows and sheep, induces a rapid and controlled luteolysis. If given as a single injection, some of the females will be in non-luteal phase or in the very early or very late luteal phase and will fail to respond. The use of two injections several days apart ensures that, at the second injection, all the animals will be at the correct stage of the cycle to respond by exhibiting oestrus behaviour and ovulation. On the other hand, progestative hormones are applied during several days (either by daily supply or using systems for a slow release) for mimicking the secretory activity of a corpus luteum during the luteal phase; withdrawal of the hormones would induce a follicular phase and, thus, oestrus and ovulation. To be effective, the duration of the progestative treatment generally has to surpass the active life of a possible corpus luteum in the ovary or may be combined with a luteolytic agent.
Synchronised oestrus is also used in laboratory animal breeding programmes for convenience and economic reasons, since mating will only occur if the recipient female is in oestrus. The oestrus cycle lasts 4-5 days in the mouse and rat (equivalent to a woman's average 28 day menstrual cycle), which leads to the need to rely on a large pool of potential recipient females to take part in potential matings. Typically, 75% of rat/mice recipients are not in oestrus in randomly cycling populations, leading to large numbers of females in a starting pool. The chance of females being in oestrus (sexually receptive, in the absence of synchronised oestrus) at the right time is 1:4 to 1:5 due to the length of their cycle. Thus, if 4 recipients are required, 16 to 20 females will be caged as potential mating pairs with 16 to 20 males, which translates to a 20-25% success rate. This figure can be even lower as some females will refuse to mate with their partner. It is known from the prior art that, in mice for example, that pregnant mare serum gonadotrophin (PMSG) at 5.0 iu (or some other agent with follicle stimulating properties such as human menopausal gonadotrophin, hMG) when administered intraperitoneally acts as a trigger and can induce hyperstimulation. This regime achieves the same goal in 46-52 h by overriding the females' endogenous hypothalamo-pituitary-ovarian axis. Thereafter, 5.0 iu human chorionic gonadotrophin (hCG) administered by the same route can be used as an adjunct ovulation trigger. Although the success rates with this approach are relatively high, it involves subjecting females to a regulated procedure under the Animals (Scientific Procedures) Act 1986 (i.e. one potentially causing pain, distress or lasting harm) and requires a skilled operator. However, the most common existing strategy for oestrus synchronisation in rodents is through the “Whitten effect”. In mice, this is achieved by exposing the target female animals to fresh soiled male cage bedding which will emanate male pheromones, including the ones produced by male preputial glands. Over the course of three days, this results in a proportion of the exposed female mice to come into oestrus. The efficacy of this strategy varies across units, housing arrangements, strains and individual animal characteristics, and is generally accepted to generate 40-70% synchronised females for use in timed mating experiments or as recipients for embryo transfer. A problem associated with this approach is the large variation in success rates and the requirement for a large pool of females to be used to in order to generate a guaranteed discrete sufficiently large number of oestrus females available for end users.
Ovulation induction and superovulation are terms to describe the use of injectable fertility drugs (gonadotrophins) to stimulate the ovaries to produce mature oocytes. The aim of ovulation induction is to grow and ovulate an oocyte in a female which would not normally ovulate at this point in time, whilst the aim of superovulation is to produce more than one oocyte to improve fertility in non-human mammalian females. In laboratory animals, superovulation, typically but not exclusively, is for the purposes of generating multiple oocytes for in vivo/in vitro fertilisation for basic research purposes or for creating transgenic/micromanipulated embryos and is achieved using well-established approaches. These typically involve administering pregnant mare serum gonadotrophin (PMSG), human menopausal gonadotrophin (hMG) or follicle stimulating hormone (FSH; recombinant or other) intraperitoneally. 46-52 h later (in rodents), the administration of a luteinising agent (hCG, luteinising hormone (LH) or other) can be used to trigger ovulation. A typical protocol for inducing superovulation in mice comprises the intraperitoneal administration of 5 iu PMSG and 5 iu hCG circa 48 h apart. These agents are absorbed by the peritoneum and recruit multiple follicles to develop to the pre-ovulatory stage (and to ovulation if given an adequate LH-like trigger), thus overriding the physiological hypothalamo-pituitary-ovarian control. As a result, more oocytes/follicles are produced per animal, offering the benefit of reducing the number of animals required to generate a given number of synchronous follicles or oocytes post-ovulation. A disadvantage to this approach is that it involves subjecting females to a regulated procedure (i.e. one potentially causing pain, distress or lasting harm) and requires a skilled operator in order to obviate complications such as accidental intravesical injection.
There is a need for a reliable method for oestrus synchronization, with fertile ovulation and mating, in a group of animals, in a short time-frame. This would be advantageous and of benefit for improving animal management (controlling and planning the reproduction of different individuals and groups in the animal units, in the presence or absence of superovulatory effects), biotechnology (artificial insemination, combination with gonadotrophin superovulatory protocols for embryo recovery and induction of pseudopregnant fosters for embryo transfer), research (scheduling standardized and consistent experimental groups with synchronized mates for generation of synchronized pregnancies either for stage-specific studies on embryonic development or for obtaining synchronized deliveries in studies on neonates) and wellbeing (reducing the number of experimental animals and refining animal-based study experimental plans).